The present application claims priority to the Chinese patent application entitled xe2x80x9cApparatus For Switching and Manipulating Particles and Method of Use Thereofxe2x80x9d, Serial No. 001290436, (NTD Patent and Trademark Agency Limited), filed on Sep. 27, 2000 the disclosure of which is incorporated herein by reference in its entirety.
1. Field of the Invention
The invention is concerned with the manipulation of particles, and more particularly, with the manipulation of small particles (e.g., cells, microbeads) using electric fields.
2. Description of the Related Art
The manipulation of particles, especially biological material, can be used to advantage in a variety of biomedical applications. The ability to manipulate individual cancer cells is of particular significance, permitting the researcher to study the interaction of either a single cancer cell or a collection of cancer cells with selected drugs in a carefully controlled environment. Various kinds of forces can be used to manipulate particles, including optical, ultrasonic, mechanical, and hydrodynamic. For example, flow cytometry has been successfully used to sort and characterize cells. Another example is the centrifuge, which has been widely used in laboratories for processing biological samples.
A current trend in the biological and biomedical sciences is the automation and miniaturization of bioanalytical devices. The development of so-called biochip-based microfluidic technologies has been of particular interest. A biochip includes a solid substrate having a surface on which biological, biochemical, and chemical reactions and processes can take place. The substrate may be thin in one dimension and may have a cross-section defined by the other dimensions in the shape of, for example, a rectangle, a circle, an ellipse, or other shapes. A biochip may also include other structures, such as, for example, channels, wells, and electrode elements, which may be incorporated into or fabricated on the substrate for facilitating biological/biochemical/chemical reactions or processes on the substrate. An important goal for researchers has been to develop fully automated and integrated devices that can perform a series of biological and biochemical reactions and procedures. Ideally, such an integrated device should be capable of processing a crude, original biological sample (e.g., blood or urine) by separating and isolating certain particles or bio-particles from the rest of the sample (e.g., cancer cells in blood, fetal nucleated cells in maternal blood, or certain types of bacteria in urine). The isolated particles are then further processed to obtain cellular components (e.g., target cells are lysed to release biomolecules, such as DNA, mRNA and protein molecules). The cellular components of interest are then isolated and processed (e.g., DNA molecules are separated and target sequences are amplified through polymerase-chain-reactions, PCR). Finally, a detection procedure is performed to detect, measure and/or quantify certain reaction products (e.g., a hybridization may be performed on the PCR-amplified DNA segments with fluorescent detection then being used to detect the hybridization result). Clearly, the ability of a biochip to manipulate and process various types of particles, including cells and cellular components from a particle mixture, would be of great significance.
Limited progress has been made to date in the manipulation of particles or bioparticles on a chip. Electronic hybridization technologies have been developed in which charged DNA molecules are manipulated and transported on an electronic chip (e.g., xe2x80x9cRapid Determination of Single Base Mismatch Mutations in DNA Hybrids by Direct Electric Field Controlxe2x80x9d, Sosnowski, R., et al., Proc. Natl. Acad. Sci., Volume 94, pages 1119-1123, 1997; xe2x80x9cElectric Field Directed Nucleic Acid Hybridization on Microchipsxe2x80x9d, Edman, C., Nucl. Acids Res., 25: pages 4907-4914, 1998, the disclosures of which are incorporated herein by reference in their entireties). Also, electrokinetic pumping and separation technologies have been developed in which biomolecules or other particles can be transported, manipulated, and separated through the use of electroosmosis and electrophoresis based kinetic effects (e.g., xe2x80x9cMicromachining a miniaturized capillary electrophoresis-based chemical analysis system on a chipxe2x80x9d, Harrison, D. J. et al, Science, Volume 261, pages: 895-896, 1993; xe2x80x9cHigh-speed separation of antisense of ligonucleotides on a micro machined capillary electrophoresis devicexe2x80x9d, Effenhauser, C. S. et al., Anal. Chem. Volume 66, pages: 2949-2953, 1994, the disclosures of which are incorporated herein by reference in their entireties). However, each of these devices suffer from limitations. Accordingly, there is a need for improved particle manipulation devices.
The present invention relates to the manipulation of particles (including bioparticles such as cells, cell organelles) using traveling-wave dielectrophoresis. The devices and methods of the present invention are suitable for the selective processing and manipulation of one kind of particle in a particle mixture. The devices and methods of the present invention are also capable of concentrating and mixing different types of particles. In addition, the devices and methods of the present invention allow the flexible and easy manipulation and control of a single type or multiple types of particles.
In one embodiment, there is provided a device for producing traveling wave electric fields, which comprises at least three electrically independent branches. Each of the branches comprises a plurality of electrodes capable of producing a traveling wave electric field in its respective branch when the electrodes in its respective branch are connected to out-of-phase signals. The branches meet at a common junction.
In another embodiment, there is provided a device for producing traveling wave electric fields, which comprises at least three sets of electrodes. The sets of electrodes are capable of producing respective traveling wave electric fields in regions adjacent to the sets of electrodes when the electrodes are connected to out-of-phase signals. The sets of electrodes are electrically independent of each other and meet at a common junction.
In another embodiment, there is provided a device for manipulating particles, which comprises at least three sets of electrodes that are electrically independent from each other. The sets of electrodes are capable of generating respective traveling-wave dielectrophoresis (twDEP) forces on particles to move the particles along respective branches when the electrodes in the sets of electrodes are connected to out-of-phase signals, and the branches are interconnected at a common junction to permit the twDEP forces to route particles from one of the branches to another of the branches. In one preferred embodiment of the device, there are three sets of electrodes, and the three sets of electrodes are oriented at about 120 degrees with respect to each other. In another preferred embodiment of the device, there are four sets of electrodes, and the four sets of electrodes are oriented at about 90 degrees with respect to each other. In yet another preferred embodiment of the device, each of the sets of electrodes comprises at least three electrodes. In another preferred embodiment of the device, the device further comprises input tubing which is in fluid communication with a particle source and at least a first one of the sets of electrodes, and may further comprise output tubing in fluid communication with at least a second one of the sets of electrodes (in which the output tubing is in fluid communication with an output reservoir). In one preferred embodiment of the device, the at least three sets of electrodes are disposed on a solid substrate, and the substrate may be selected from the group consisting of silicon, glass, ceramics, and plastics. In another preferred embodiment of the device, the device further comprises a substrate on which the sets of electrodes are disposed, a cover having at least one port therein through which the input tubing passes, and a spacer element disposed between the substrate and the cover, in which the spacer element has an opening therein through which the particles to be manipulated are introduced from the input tubing. In one preferred embodiment of the device, the device further comprises at least one electrical signal source which applies AC voltages to each electrode in the sets of electrodes, in which the phases of the voltages applied to the electrodes of the sets of electrodes are selected to induce respective traveling wave electric fields along the branches. For example, these phases may be at 0, 90, 180, and 270 degrees with respect to each other. In another preferred embodiment of the device, the device further comprises conductor elements that extend from the electrodes to connection pads, in which the connection pads are connected to at least one signal generator. The electrodes in any given one of the sets of electrodes may all be connected to different connection pads, or alternatively, in any given one of the sets of electrodes, adjacent electrodes are connected to different connection pads and more than one electrode is connected to at least one of the connection pads. In a preferred embodiment of the device, the voltages applied to adjacent electrodes are out-of-phase with each other. In yet another preferred embodiment of the device, the device further comprises an electrically-independent, linear electrode set located adjacent one of the sets of electrodes, in which the linear electrode set is capable of producing traveling wave electric fields. In a preferred embodiment of the device, the particles comprise biological material. The biological material may comprise at least one member selected from the group consisting of cells, organelles, cell aggregates, biomolecule-covered microparticles, and complexes between moieties and their binding partners. Alternatively, the particles may comprise non-biological material. In yet another preferred embodiment of the device, at least one electrode that is disposed near the common junction has a curvature therein. In another preferred embodiment of the device, the electrodes have a configuration which is generally pointed. In yet another preferred embodiment of the device, the electrodes in each of the sets of electrodes are concentric arc segments with decreasing size towards the common junction.
In another embodiment, there is provided a device for manipulating particles, which comprises a first set of electrodes capable of generating traveling wave dielectrophoresis (twDEP) forces on particles which move the particles along a branch when the electrodes in the first set of electrodes are connected to out-of-phase signals. The device also comprises a second set of electrodes capable of generating a force, which urges the particles toward the center of the branch when the electrodes in the second set of electrodes are connected to at least one electrical signal. In a preferred embodiment, the first set of electrodes is disposed on a first substrate, the second set of electrodes is disposed on a second substrate, and the substrates are separated by a spacer.
In yet another embodiment, there is provided a device for manipulating particles, which comprises at least three branches. Each branch comprises a first set of electrodes capable of generating traveling wave dielectrophoresis (twDEP) forces on particles, which move the particles along a branch when the electrodes in the first set of electrodes are connected to out-of-phase signals. The branch also comprises a second set of electrodes capable of generating a force, which urges the particles toward the center of the branch when the electrodes in the second set of electrodes are connected to signals. The at least three branches meet at a common junction to permit the twDEP forces to route particles from one of the branches to another of the branches. In a preferred embodiment of the device, the second set of electrodes generate conventional dielectrophoresis (cDEP) forces, and the electrodes of the second set are oriented substantially perpendicular to the electrodes of the first set.
In another embodiment, there is provided a device for manipulating particles, which comprises an array of devices connected to one another. Each of the devices in the array comprises at least three sets of electrodes, which are electrically independent from each other. The sets of electrodes are capable of generating respective traveling-wave dielectrophoresis (twDEP) forces on particles to move the particles along respective branches when the electrodes in the sets of electrodes are connected to out-of-phase signals, and the branches are interconnected at a common junction to permit the twDEP forces to route particles from one of the branches to another of the branches.
In yet another embodiment, there is provided a method of transporting particles. The method comprises providing a plurality of electrodes, which are spaced apart from each other; and applying a voltage of a first polarity to a first electrode to attract particles having a net charge of a polarity opposite to the first polarity. The method comprises transporting the particles to a second electrode by applying a voltage of the first polarity to the second electrode, while reducing the magnitude of the voltage applied to the first electrode, so that the charged particles are moved away from the first electrode and attracted towards the second electrode; this transporting procedure is repeated for other electrodes to transport the particles from one electrode to another electrode in a step-wise fashion. In one preferred embodiment of the method, the plurality of electrodes comprises at least three sets of electrodes capable of generating forces on charged particles for moving the charged particles along respective branches, wherein the sets of electrodes are electrically independent from each other and the branches are interconnected at a common junction to permit the forces to route charged particles from one of the branches to another of the branches. In yet another preferred embodiment of the method, the step of reducing the magnitude of the voltage comprises reversing polarity of the first electrode. In another preferred embodiment of the method, the transporting step comprises applying a voltage of a polarity opposite to the first polarity to all the electrodes except the second electrode.
In another embodiment, there is provided a method of sorting particles. The method comprises providing a device that comprises at least three sets of electrodes which are electrically independent from each other, in which the sets of electrodes are capable of generating respective traveling-wave dielectrophoresis (twDEP) forces on particles to move the particles along respective branches when the electrodes in the sets of electrodes are connected to out-of-phase signals, and the branches are interconnected at a common junction to permit the twDEP forces to route particles from one of the branches to another of the branches. The method comprises introducing a sample comprising at least two types of particles into the device, and generating traveling wave dielectrophoresis forces at the junction such that at least one of the particle types travels away from the junction in a first direction and at least one of the other particle types travels away from the junction in a second direction. In a preferred embodiment of the method, the twDEP forces are generated by applying voltages to the sets of electrodes. In yet another preferred embodiment of the method, the method further comprises identifying at least one particle type before the particle type enters the junction and applying voltages to the sets of electrodes, in which the voltages are selected in view of the result of the identifying step. In yet another preferred embodiment of the method, the identifying step comprises monitoring the fluorescence of the particle types.
In another embodiment, there is provided a method of combining different kinds of particles, which comprises introducing at least a first kind of particle into a first branch of a device comprising at least three sets of electrodes which are electrically independent from each other. The sets of electrodes in the device are capable of generating respective traveling-wave dielectrophoresis (twDEP) forces on particles to move the particles along respective branches when the electrodes in the sets of electrodes are connected to out-of-phase signals, and the branches are interconnected at a common junction to permit the twDEP forces to route particles from one of the branches to another of the branches. The method comprises introducing at least a second kind of particle into a second branch of the device, and transporting the at least first kind and at least the second kind of particles towards the junction such that the at least first kind of particle and the at least second kind of particle are combined with one another at the junction.
In another embodiment, there is provided a method of concentrating particles, which comprises introducing particles into at least first and second branches of a device. The device comprises at least three sets of electrodes which are electrically independent from each other. The sets of electrodes are capable of generating respective traveling-wave dielectrophoresis (twDEP) forces on particles to move the particles along respective branches when the electrodes in the sets of electrodes are connected to out-of-phase signals, and the branches are interconnected at a common junction to permit the twDEP forces to route particles from one of the branches to another of the branches. The method comprises transporting the particles towards the junction such that the particles are concentrated at the junction. In one preferred embodiment of the method, the method comprises introducing particles into at least three branches of the device, and transporting the particles in the at least three branches towards the junction such that the particles are concentrated at the junction.
In yet another embodiment, there is provided a method of dispersing particles, which comprises introducing particles into a common junction of a device. The device comprises at least three sets of electrodes which are electrically independent from each other. The sets of electrodes are capable of generating respective traveling-wave dielectrophoresis (twDEP) forces on particles to move the particles along respective branches when the electrodes in the sets of electrodes are connected to out-of-phase signals, and the branches are interconnected at a common junction to permit the twDEP forces to route particles from one of the branches to another of the branches. The method comprises dispersing the particles away from the junction and into at least two of the branches. In a preferred embodiment of the method, the method comprises dispersing the particles away from the junction and into three branches.
In another embodiment, there is provided a method of separating a first kind of particle from a second kind of particle, the first and second kinds of particle being distributed throughout a device. The device comprises at least three sets of electrodes, which are electrically independent from each other. The sets of electrodes are capable of generating respective traveling-wave dielectrophoresis (twDEP) forces on particles to move the particles along respective branches when the electrodes in the sets of electrodes are connected to out-of-phase signals, and the branches are interconnected at a common junction to permit the twDEP forces to route particles from one of the branches to another of the branches. The method comprises applying conventional dielectrophoresis (cDEP) forces to the first kind of particle to cause the first kind of particle to remain stationary, and applying traveling wave dielectrophoresis (twDEP) forces to the second kind of particle to cause the second kind of particle to be moved away from and thereby be separated from the first kind of particle. In a preferred embodiment of the method, the method comprises causing the first kind of particle to be attracted to electrodes where the first kind of particle is held in place by the cDEP forces. In another preferred embodiment of the method, the method comprises diverting the second kind of particle to a desired branch. In yet another preferred embodiment of the method, the separation comprises applying electric fields of different frequencies to the electrodes, in which the fields of the different frequencies interact differently with the first and second kinds of particle to cause the first kind of particle to remain stationary while the second kind of particle is transported. In yet another preferred embodiment of the method, first and second kinds of particle are introduced onto electrodes in the form of a particle suspension. In still another preferred embodiment of the method, the method comprises separating out a third kind of particle from the first and second kinds of particle.
In another embodiment, there is provided a method of separating a first kind of particle from a second kind of particle, which comprises introducing a continuous stream of fluid having the first and second kinds of particle in suspension into a device. The device comprises at least three sets of electrodes, which are electrically independent from each other. The sets of electrodes are capable of generating respective traveling-wave dielectrophoresis (twDEP) forces on particles to move the particles along respective branches when the electrodes in the sets of electrodes are connected to out-of-phase signals, and the branches are interconnected at a common junction to permit the twDEP forces to route particles from one of the branches to another of the branches. The method also comprises applying conventional dielectrophoresis (cDEP) forces to the first kind of particle to cause the first kind of particle to be attracted to electrodes where the first kind of particle is held in place by cDEP forces while the second kind of particle is carried with the stream of fluid and thereby separated from the first kind of particle. The method comprises ceasing the stream of fluid, and applying electric voltages to the electrodes at a frequency selected to generate traveling wave dielectrophoresis forces for transporting the first kind of particle.
In yet another embodiment, there is provided a method of separating a first kind of particle from a second kind of particle, which comprises introducing a continuous stream of fluid having first and second kinds of particle in suspension into a device. The device comprises at least three sets of electrodes which are electrically independent from each other. The sets of electrodes are capable of generating respective traveling-wave dielectrophoresis (twDEP) forces on particles to move the particles along respective branches when the electrodes in the sets of electrodes are connected to out-of-phase signals, and the branches are interconnected at a common junction to permit the twDEP forces to route particles from one of the branches to another of the branches. The device comprises an electrically-independent, linear electrode set located adjacent one of the sets of electrodes, in which the linear electrode set is capable of producing traveling wave electric fields. The method comprises applying electric voltages to the linear electrode set to produce conventional dielectrophoresis (cDEP) forces on the first and second kinds of particles so that the first and second kinds of particles are attracted to electrodes of the linear electrode set where the first and second kinds of particles are held in place by the cDEP forces. The method comprises ceasing the stream of fluid, and applying electric voltages to the linear electrode set and to the at least three sets of electrodes at frequencies and phases selected to generate traveling wave dielectrophoresis forces for transporting the first kind of particle to the end of one branch and transporting the second kind of particles to the end of another branch.